The present invention concerns a new growth/differentiation factor of the TGF-xcex2 family and DNA sequences coding therefor.
The TGF-xcex2 family of growth factors which includes BMP-, TGF- and inhibin-related proteins (Roberts and Sporn, Handbook of Experimental Pharmacology 95 (1990), 419-472) is particularly relevant for a wide range of medical treatment methods and applications. These factors are suitable in methods which concern wound-healing and tissue regeneration. Furthermore several members from the TGF-xcex2 family induce tissue growth, in particular growth of bones, and therefore play a crucial role in inducing the development of cartilage and bones.
Wozney (Progress in Growth Factor Research 1 (1989), 267-280) and Vale et al (Handbook of Experimental Pharmacology 95 (1990), 211-248) describe various growth factors such as those which are related to the BMP group (bone morphogenetic proteins) and the inhibin group. The members of these groups show significant structural similarities. The precursor of the protein consists of an amino-terminal signal sequence, a propeptide and a carboxy-terminal sequence of about 110 amino acids that are cleaved from the precursor and constitute the mature protein. In addition their members are defined by an amino acid sequence homology. The mature protein contains the most conserved sequences, in particular seven cysteine residues which are conserved among the family members. The TGF-xcex2-like proteins are multifunctional, hormonally active growth factors. They also have related biological activities such as chemotactic attraction of cells, promotion of cell differentiation and tissue-inducing capabilities such as cartilage-inducing and bone-inducing capabilities. The U.S. Pat. No. 5,013,649 discloses DNA sequences that code for osteo-inductive proteins that are denoted BMP-2 and the U.S. patent applications Ser. No. 179 101 and 170 197 disclose the BMP proteins BMP-1 and BMP-3. Moreover many types of cells are able to synthesize TGF-xcex2-like proteins and practically all cells have TGF-xcex2 receptors.
On the whole these proteins show differences in their structure which leads to significant variations in their exact biological function. In addition they are found in a wide range of different types of tissue and at various stages of development. As a result they can exhibit differences with regard to their exact function e.g. the required cellular physiological environment, their life-span, their target sites, their requirements for auxiliary factors and their stability against degradation. Thus, although a multitude of proteins have been described that exhibit a tissue-inductive and in particular osteo-inductive potential, their natural functions in the organism andxe2x80x94more significantlyxe2x80x94their medical relevance still have to be investigated in detail. It is thought to be highly probable that members of the TGF-xcex2 family are present that are still unknown which are important for osteogenesis or the differentiation/induction of other types of tissue. A major difficulty in the isolation of these new TGF-xcex2-like proteins is, however, that their functions cannot yet be described exactly enough to develop highly discriminating bioassays. On the other hand the expected nucleotide sequence homology to other members of the family is too low to enable a screening by classical nucleic acid hybridization techniques. Nevertheless the further isolation and characterization of new TGF-xcex2-like proteins is urgently required in order to provide further inducing and differentiation-promoting proteins that fulfil all the desired medical requirements. These factors could be used medically in the healing of lesions and the treatment of degenerative diseases of bones and/or other types of tissue such as the kidney or the liver.
A nucleotide and amino acid sequence for the TGF-xcex2 protein MP-52 is stated in the Patent Application PCT/EP93/00350 in which the sequence corresponding to the mature peptide and a major portion of the sequence corresponding to the propeptide of MP-52 are given. The complete sequence of the propeptide MP-52 is not disclosed.
The object on which the present invention is based is to provide DNA sequences that code for new members of the TGF-xcex2 protein family with mitogenic and/or differentiation-inductive e.g. osteo-inductive potential. The object of the present invention was therefore in particular to provide the complete DNA and amino acid sequence of the TGF protein MP-52.
This object is achieved by a DNA molecule that codes for a protein of the TGF-xcex2 family and which comprises
(a) the part coding for the mature protein and if desired further functional parts of the nucleotide sequence shown in SEQ ID NO. 1,
(b) a nucleotide sequence corresponding to a sequence from (a) within the scope of the degeneracy of the genetic code,
(c) an allelic derivative of a nucleotide sequence corresponding to one of the sequences from (a) and (b) or
(d) a sequence hybridizing with one of the sequences from (a), (b) or (c) provided that a DNA molecule according to (d) contains at least the part coding for a mature protein from the TGF-xcex2 family.
Further embodiments of the present invention concern the subject matter of the further claims. Other features and advantages of the invention can be derived from the description of the preferred embodiments and figures. The sequence protocols and figures are now briefly described.
SEQ ID NO. 1 shows the complete nucleotide sequence of the DNA coding for the TGF-xcex2 protein MP-52. The ATG start codon starts with nucleotide 640. The start of the mature protein begins after nucleotide 1782.
SEQ ID NO. 2 shows the complete amino acid sequence of the TGF-xcex2 protein MP-52 which was derived from the nucleotide sequence shown in SEQ ID NO. 1.